本文選題：蘇云金芽孢桿菌 + vip基因　； 參考：《江蘇農(nóng)業(yè)科學(xué)》2017年04期

【摘要】：營養(yǎng)期殺蟲蛋白(vegetative insecticidal protein,簡稱VIP)是蘇云金芽孢桿菌(Bacillus thuringiensis,Bt)在營養(yǎng)期表達(dá)的一類新型殺蟲毒蛋白,該蛋白的發(fā)現(xiàn)使得Bt殺蟲劑在殺蟲活性、殺蟲范圍方面得到很大突破。首次比較我國不同氣候類型(熱帶:海南省,亞熱帶:廣西壯族自治區(qū),溫帶:黑龍江省)的Bt菌株和vip基因的分布情況。結(jié)果表明,從海南省841份土樣中分離出156株Bt菌(18.55%),從廣西地區(qū)1 420份土樣中分離出356株Bt菌(25.07%),從黑龍江省1 010份土樣中分離出167株Bt菌(16.53%);從海南省的156株Bt菌中鑒定出22個vip基因(占14.1%),從廣西地區(qū)的356株Bt菌中鑒定出2個vip基因(占5.62%),然而,從黑龍江省的156株Bt菌中沒有鑒定出vip基因(0%)。利用聚合酶鏈?zhǔn)椒磻?yīng)(polymerase chain reaction,簡稱PCR)方法成功克隆出1個vip3Aa的全長基因并獲得登錄號KT792883,將該基因插入到表達(dá)載體p ET-21b中,轉(zhuǎn)化菌株Rosetta(DE3),在低溫條件下表達(dá)88 ku的蛋白,將該蛋白進(jìn)行生物活性測定,結(jié)果顯示,該蛋白對甜菜夜蛾(Spodoptera exigua)具有較好的殺蟲活性,LC50為5.124 ng/mg,對棉鈴蟲(Heliothis armigera)活性較低,LC50為870.1 ng/mg。研究結(jié)果顯示,Bt菌株和vip基因的分布有地域性差異,在溫度較高的熱帶、亞熱帶地區(qū)有比較豐富的Bt菌株和vip基因資源。研究將有利于充分利用土壤中的Bt菌株,分離具有自主知識產(chǎn)權(quán)和重要應(yīng)用價值的新型vip抗蟲基因,這將會為研究新型殺蟲蛋白、延緩害蟲抗性問題提供有益幫助。
[Abstract]:Vegetative insecticidal protein (vip) is a new type of insecticidal protein expressed by Bacillus thuringiensis Bt. during the nutritive period. The discovery of this protein has made a great breakthrough in insecticidal activity and scope of BT insecticide. It is the first time to compare the distribution of BT strain and vip gene in different climatic types of China (tropical: Hainan province, subtropical region: Guangxi Zhuang Autonomous region, temperate zone: Heilongjiang Province). The results show that 156 strains of BT bacteria were isolated from 841 soil samples in Hainan Province, 356 strains of BT bacteria were isolated from 1,420 soil samples from Guangxi region, 167 strains of BT bacteria were isolated from 1,010 soil samples in Heilongjiang Province, and 22 strains were identified from 156 strains of BT bacteria in Hainan Province. Two vip genes (5.62%) were identified from 356 BT strains in Guangxi. The vip gene was not identified from 156 BT strains in Heilongjiang Province. A full-length vip3Aa gene was successfully cloned by polymerase chain reaction (PCR) method, and the accession number KT792883 was obtained. The gene was inserted into the expression vector p ET-21b. The transformed strain Rosetta-DE3H expressed 88 ku protein at low temperature. The bioactivity of the protein was determined. The results showed that the protein had a better insecticidal activity against Spodoptera exigua (5.124 ng / mg) and a lower activity of 870.1 ng / mgagainst Helicoverpa armigera (Heliothis armigera). The results showed that there were regional differences in the distribution of BT and vip genes, and there were abundant BT strains and vip gene resources in the tropics and subtropics where the temperature was higher. The study will be beneficial to the utilization of BT strains in soil and the isolation of new vip insecticidal genes with independent intellectual property rights and important application value, which will provide useful help for studying new insecticidal proteins and delaying the problem of pest resistance.
【作者單位】： 東北農(nóng)業(yè)大學(xué);
【基金】：國家自然科學(xué)基金(編號:31401812) 東北農(nóng)業(yè)大學(xué)科學(xué)研究基金(編號:2010RCB54)
【分類號】：S476

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